Abstract
The superbacteria Aeromonas veronii displays not only a strong pathogenicity but also the resistance to nine kinds of antibiotics, resulting in the economic losses and health hazards. Small Protein B (SmpB) plays an important role in protein quality control, virulence, and stress reactions. Transcriptomic data revealed that expressions of the type IV pilus assembly and type VI secretion system (T6SS) proteins were downregulated in SmpB deficiency, indicating that the virulence of A. veronii might be attenuated. Although SmpB deletion decreased colonization in the mouse spleen and liver, LD50 of the smpB mutant was not altered as expected, compared with the wild type. Further, the transcriptomic and quantitative RT-PCR analyses showed that the combination of the downregulated AvrA and the upregulated iron-sulfur protein activator IscR, mediated the oxidative tolerance in smpB deletion. Next a reporter plasmid was constructed in which the promoter of iscR was applied to control the expression of the enhanced green fluorescent protein (eGFP) gene. When the reporter plasmid was co-expressed with the AvrA expression into E. coli, the relative fluorescence intensity was decreased significantly, suggesting that AvrA bound to iscR mRNA by base pairing, which in turn relieved the inhibition of iscR and intensified the downstream iron-sulfur proteins. Collectively, the smpB mutant exhibited an attenuated virulence in mice and enhanced tolerances to oxidative stress. This study demonstrates the complexity of gene regulation networks mediated by sRNA in systems biology, and also reflects the strong adaptability of superbacteria A. veronii in the process of evolution.
Highlights
With the application of antimicrobial drugs, a wide range of mechanisms have evolved for bacteria to combat the huge selection pressures of antibacterial agents
To determine the differences in pathways and gene expressions between wild type and smpB knockout in the M9 medium, the transcriptome assembly of A. veronii was generated from 91.17 million paired-end RNA-seq reads using Illumina Xten technology and a total of 22,055 transcripts were mapped by genome NZ_CP012504.1 (Figure 1A)
Sixteen pathways were significantly affected by Small Protein B (SmpB) knockout, including sulfur metabolism, biosynthesis of siderophore group non-ribosomal peptides, bacterial chemotaxis, arginine and peptidoglycan biosynthesis, and microbial metabolism in a diverse environment (Figure 1B)
Summary
With the application of antimicrobial drugs, a wide range of mechanisms have evolved for bacteria to combat the huge selection pressures of antibacterial agents. The capacities of oxidative stress and iron were increased by sRNA downregulation in smpB knockout. A sRNA designated as AvrA was revealed to negatively regulate the expression of IscR (ironsulfur cluster assembly transcriptional regulator) IscR governs iron hemostasis as a global regulator during growth and stress responses (Aguilera-Arreola et al, 2007), which exhibits a resistance to oxidants (Mandin et al, 2016), and represses ironsulfur metabolism under iron starvation (Haines et al, 2015; Carrier et al, 2017). Some bacteria employ ArfA and ArfB to rescue stalled ribosomes when smpB is deficient (Keiler, 2015; Liu et al, 2016; Huter et al, 2017). SRNA AvrA releases the trapped IscR promotor to activate the expression of iron sulfur proteins, remedying the iron deficiency after smpB destruction. In the evolutionary history of bacteria, it offers additional evidence for the formation of superbacteria
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